Method and apparatus for diagnosis, detection of cell abnormalities and morphology of living systems

ABSTRACT

A method and apparatus for dielectric diagnostic analysis of human and non-human cells or tissue functions by measuring the response of the cells or tissue to an applied excitation signal in a time period less than the polarization relaxation time period of a domain group of the cells or tissue under examination.

TECHNICAL FIELD

The present invention relates to an apparatus and method of diagnosticmeasurement and in particular, a method and apparatus adapted to analysevarious parameters of living materials and specimens to determine thedielectric characteristic of a specimen under test for the purpose ofdiagnosis of the state of the specimen. Depending on the specimen, awide range of states may be susceptible to diagnosis including such asdisease in plants, animals or humans: the revelation of residual toxinsin consumer goods from dairy products, meat products, fruit andvegetable products, fish, grains and stock feed, oils and other liquids.

The present invention further helps identify abnormalities andtransformations in living bodies in their earliest stages, much beforethe clinical appearance of a disease.

BACKGROUND ART

At the present state of technology it is well known that the dielectricbehaviour of such as plant, fruit, animal and human tissue correspondsto broad features in their composition and structure. Recent studieshave revealed that the cell is a highly ordered dynamic entity whichacts holistically with respect to chemical and physical events within aliving body, and the existence of domains in the cytoplasm is a generalrule. These domains are electrically polarised units of ordered, packedbiopolymers in "biowater". The different organs in a living organism,with compartmental similarity and harmonised metabolism, have basicdifferences in domain arrangements which lead to a difference indielectric responses. A disease transformation in a living body whichhas a viral origin or resulting from the action of toxins and otherchemicals also changes the domain structure and hence the polarisationand dielectric response of the tissue or cell.

A domain is herein defined as a region of a system, or a region of asubstance, comprising atoms or molecules which can be thought of as asingle entity; this single entity being responsive to electric ormagnetic fields and includes such a system having a plurality of theseentities. Examples of a domain include; a ferroelectric or ferromagneticdomain, a cluster of atoms or molecules, an organic cell, a bacterium, avirus, a cluster or collection of cells.

A domain group is a collection of said domains having the same responseto an electric or magnetic field.

In the past, precise measurement of parameters of domains wereinconceivable due to limitations of the instruments. Measurements ofrelative dielectric permittivity, energy dissipation and electricalimpedance are not possible due to very high values of electricalconductance overshadowing real kinetic characteristics. Existing methodsof measurement are mostly based on impedance bridges, which areinadequate at frequencies below 100 Hz due to noise instability,electrode polarisation and the time required to obtain balancedconditions. These bridges yield relative permittivity, energydissipation and electrical impedance values only at discrete frequenciesand therefore each frequency setting causes disruption of sequentialmeasurements. The dielectric properties of living tissue from bodieswill change when they are taken out of their natural environment. Deadtissue will show a greater change with changes of cell morphology.Conductivity measurement is mostly carried out by D.C. electrometers ofwide current range, often from 10⁻¹⁴ Ampere to a few milliAmpere. Thisrange being covered by switching to sequential decade ranges with amismatch of measured current values. A.C. and D.C. measurements requiredifferent apparatus, separate sample settings and long time switchingintervals from one instrument to the next. The morphological changes ofa cell are much faster, so the obtained parameters will refer todifferent intracellular structures resulting in an incorrect correlationbetween these parameters. Sample size limitations sometimes up to a fewmilligrams reduces electrode sensitivity and field noise overshadows theresults for fine structural studies.

DISCLOSURE OF INVENTION

In an effort to ameliorate the disadvantages of the prior art or atleast to provide a commercially viable alternative to the prior art, thepresent invention proposes a dielectric diagnostic analyser (DDA) and amethod of diagnosis.

In a first aspect, the present invention consists in an apparatusadapted to perform diagnostic analysis of a specimen having at least onedomain group as hereinbefore defined, the apparatus comprising:

excitation generating means to generate a predetermined excitationsignal;

measuring means to measure a response signal of the specimen to thepredetermined excitation signal:

electrode means for transmitting and receiving the predeterminedexcitation signal and response signal of the specimen, respectively;

analysing means arranged to analyse said response signal; and

switching means adapted to switch the electrode means between themeasuring means, and excitation generating means, in a time period lessthan a polarization relaxation time period of the at least one domaingroup in the specimen.

Preferably, the excitation generating means is the source of thepredetermined excitation signal and may be an electrometer or afrequency bridge adapted to generate a predetermined signal. In one formof the invention the measuring means compares an electrometer or afrequency bridge arranged to measure responses received at the electrodemeans as the response signals of the system.

Typically the analysing means comprises an electronic computer,electrometer and frequency bridge arranged to analyse the responsesignals, received at the electrode means and the computer has a displayfor displaying a diagnostic result. Preferably the switching means isalso controlled by the computer which allows switching of the electrodemeans between the excitation means and the measuring means at times lessthan the smallest relaxation time, of the polarized domain group, to bemeasured.

In an embodiment of this invention the electrode means is in the form ofa suction cup electrode, a pinch electrode, a thermocontrolled electrodeor any combination of two or more similar electrodes.

In a second aspect, the present invention provides a method ofdiagnostic analysis comprising;

applying a predetermined first excitation signal to a specimen having atleast one domain group, as hereinbefore defined, so as to elicit aresponse from the domain group within the specimen;

analysing the response from the domain group to determine the maximumresponse of each domain group; and

comparing said maximum response to a maximum response of a controlspecimen.

Preferably the first excitation signal is a ramp function voltage sweepor a time rate of change of voltage, and the response from the domain,in the domain group of the specimen, is measured as a change in acurrent flow through the specimen over time.

Typically the point of maximum response is at the threshold polarizationvoltage of each domain group and is representative of a maxima in thepolarization of each domain group of the specimen.

Preferably a control specimen is any specimen, analogous to the specimento be diagnosed and considered to be the statistical norm of thatspecimen.

In an alternative form of the second aspect of the present invention,the first excitation signal is a frequency dependent applied voltage andthe response from the domains is measured so as to allow thedetermination of dielectric permittivity, and dissipation energy, ofeach domain group. In this form the point of maximum response of eachdomain group is determined by a local maxima in the dielectricpermittivity or a local minima in the dissipation energy of that domaingroup.

In a third aspect, the present invention provides a method of diagnosticanalysis comprising:

all the steps of the second aspect of the present invention as well as;

applying a second excitation signal corresponding to a signal value at,or near, the point of maximum response of each domain group to elicit afurther response in each domain group; and

detecting the variation and length of said further response upon removalof the second excitation signal.

Preferably the second excitation signal is applied in the absence of thefirst excitation signal, and the further response is measured uponremoval of the second excitation signal while each domain is relaxing toits natural state.

Typically the detecting of the variation and length of the furtherresponse occurs within the time in which the domains in each group relaxto the state they were in before the second excitation signal wasapplied.

BRIEF DESCRIPTION OF DRAWINGS

The present invention will now be described by way of example only, withreference to the accompanying drawings, in which:

FIG. 1 is a schematic diagram of a dielectric diagnostic analyser inaccord with an embodiment of the invention:

FIG. 2 is a schematic diagram of a preferred embodiment of a suction cupelectrode;

FIG. 3 is a schematic diagram of a preferred embodiment of a pinchelectrode;

FIG. 4 is a schematic diagram of a preferred embodiment of athermo-controlled electrode and chamber;

FIG. 5 shows an hysteresis graph of the dielectric permittivity againstthe applied electrode pressure on living tissue;

FIG. 6 is a schematic diagram of the equivalent circuit of theinductive, capacitive and resistive processes in the dielectric responsemechanism of domain structures analysed by the method and apparatus ofthe present invention.

FIG. 7 is the printout of a computer screen of two graphs of currentversus voltage each having two plotted curves, the right hand side graphbeing an enlarged view of a section of the left hand side graph;

FIG. 8 is a graph containing 3 curves of the relative permittivityversus the frequency for a rat shown on a logarithmic-logarithmic scale.

FIG. 9 is a graph containing 3 curves of the dissipation factor versusthe frequency on logarithmic scale for a rat;

FIG. 10 is an example of discharge current versus time curve for domainstructure in a Wistar rat thigh muscle:

FIG. 11 is a table of relaxation time constants for four domains (showsas τ₁, τ₂, τ₃, τ₄) for a plurality of rat organs;

FIG. 12 is a graph of a Fourier analysis of four domains in theintracellular structure of the thigh muscle of a Wistar rat;

FIGS. 13(a) and (c) are graphs of the dielectric permittivity as afunction of frequency, while FIGS. 13(b) and (d) are graphs of thedissipation factor versus frequency: all for various temperaturesettings of the tongue tissue of a Wistar rat;

FIG. 13(e) is a graph of current versus applied voltage for tonguetissue at 20 degrees Celsius, before and after the tissue was heatedabove 42.5 degrees Celsius;

FIG. 13(f) is a graph of the frequency response of the tissues versusthe inverse of the absolute temperature (temperature measured in degreesKelvins).

FIG. 14 shows a table of dielectric properties of tissue of rat organs.

BEST MODES

FIG. 1 shows an embodiment of the first aspect of the present invention,which comprises a switching means 97 connected to a frequency bridge 95,an electrometer 96 and a computer unit 91 via appropriately shieldedcables. The computer unit 91 is also connected to a keyboard 92, adisplay monitor 93 and a printer unit 94 in the usual way to provide acomputer system. The electrometer 96 and frequency bridge 95 are alsoconnected to the computer unit 91, such that an operator can through theuse of the keyboard 92 instruct the computer unit 91 to change thesettings on the electrometer 96 or the frequency bridge 95. Preferably,the electrometer 96 and the frequency bridge 95 has the additionaloption of changing the settings manually. The computer unit 91 can beprogrammed to receive input signals, from the electrometer 96 and thefrequency bridge 95, which can be analysed by means of dedicatedsoftware programmes such as Intel's IEEE 488 and then to output theresulting analysis on the display monitor 93 or printer 94.

The switching means 97 further having connections via a plurality ofelectrically shielded conducting cables to three electrode devices. Thecomputer unit 91 is programmed to instruct the switch means 97 to switchbetween any one of the three electrode devices. The first electrodedevice as illustrated in FIG. 1 and FIG. 2 is a suction cup electrode118 which comprises an excitation electrode 114 to induce a current in atissue specimen 117, a measuring electrode 115 to measure the responsesignals of the specimen 117 resulting from the excitation induced by theexcitation electrode 114, a guard electrode 116 to prevent unwantedsurface currents reaching the measuring electrode 115, and a suctiondevice 112 connected to the suction cup electrode 118 by way of anairflow link 113 to the air passage channel 125 of the suction cup 118.The suction device 112 is used to adjust the pressure within the suctioncup electrode 118, so that not only does the cup adhere to the specimenbut the contact pressure between the specimen 117 and the electrodes(i.e. the excitation electrode 114, the guard electrode 116 andmeasuring electrode 115) can be adjusted to an optimum pressure. Theoptimum pressure between the electrodes and specimen is obtained from alocal maximum value of the dielectric permittivity in a hysteresis plot,as shown in FIG. 5. The excitation and measuring electrodes 114 and 115,respectively, are set to the optimum pressure before diagnosticmeasurements are obtained. The second electrode device illustrated inFIG. 1 and FIG. 3 is hereinafter referred to as the pinch electrode 101which comprises an excitation electrode 100, mounted on one jaw of apair of pincers 136. while the guard 98 and measuring electrode 99 aremounted on the opposite jaw of the pair of pincers 136. At the other endof the pair of pincers 136, a spring 132, an adjusting screw mechanism131 and a micrometer measuring gauge 130 are arranged to adjust andmeasure the distance between the excitation electrode 100 and themeasuring electrode 99 at the jaw end of the pair of pincers 136. Aspecimen 102 is pinched between the electrodes at the jaw end and aforce between the jaws is applied by the adjustment of the screwmechanisms 131 and spring until the desired distance is read off thegauge reflecting the distance between the excitation electrode 98 andthe measuring electrode 99 at the jaw end sandwiching the specimen 102between the electrodes.

The third electrode device illustrated by FIG. 1 and FIG. 4 ishereinafter referred to as the thermocontrolled electrode 105 whichcomprises a first piston 141, of electrically conductive material tofunction as the excitation electrode 108, and fits within a first tefloncylinder 147 so that it protrudes from both ends. The said first tefloncylinder 147 has, a guard electrode 107 which wraps around one end ofthe outer surface of the cylinder 147 and an electromagnetic shield 144which wraps around the other end of the outer surface of the tefloncylinder 147.

A second teflon cylinder 149 substantially similar to the first tefloncylinder 147, has a second piston 142 functioning as the measuringelectrode 106. Piston 142 is allowed to slide in and out of the cylinder149 by means of an adjusting nut 145 located at the end of the piston142 which protrudes from second teflon cylinder 149 nearest to theelectromagnetic shield 144. A cap 146 placed over the nut 145 stops itfrom turning at will. The guard electrode 107 on the second tefloncylinder 149 extends beyond the end of the cylinder 149. The two tefloncylinders 147, 149 slide, with some frictional force, into a thirdcylinder so that the guard electrodes 107 meet, leaving a gap betweenthe excitation electrode 108 and the measuring electrode 106 to fit aspecimen 112. The adjusting nut 145 can then be used to change thedistance between the gap. The third cylinder being a thermocontrolledjacket 148 with two ports 150 so that fluid can be pumped in or out, ata predetermined temperature, to thermally control the specimen 112. Thethermocontrolled jacket 148 is connected to a thermocontrol unit 110(seen in FIG. 1) by means of tubing to the ports 150. The thermocontrolunit 110 being capable of adjusting the flow rate and temperature of thefluid within the jacket 148. The thermocontrol unit 110 further having afeedback cable 109 to the computer unit 91, so that the flow rate andtemperature of the fluid can be set or monitored.

The second aspect of the present invention comprises a method ofdiagnostic analysis of the human body or specimen under test. Thefollowing parameters can be measured directly, or indirectly by way ofcalculations; current, voltage, specific surface conductance, specificvolume conductance, domain relaxation time constants, capacitance,inductance, relative permittivity, impedance, reactance and dissipationfactor at different frequencies and temperatures.

FIG. 6 is a schematic diagram of the equivalent electric circuit for theresistive, capacitive and inductive processes in the intracellularmorphology based on the known concepts of domain structures.

The embodiment of apparatus of the first aspect of this inventionhereinbefore described, enables the measurement of various parameters byexciting the intracellular domains and measuring those parameters withinthe relaxation time periods of the domains to thereby ameliorate theproblem of electrode polarisation obscuring the measurements. Values ofthese parameters are therefore revealed by measuring these parametersduring the relaxation cycle after excitation.

By way of example only, we will demonstrate how the diagnostic resultsare obtained, for the induction of cancer in a Wistar rat, using thepinch electrode 101 hereinbefore described.

FIG. 7 is a printout of two graphs for the current versus voltageapplied to a Wistar rat, the right hand side graph being an enlargementof a section of the curves on the left hand side graph. The curve 301represents the results of a test on the tongue tissue of a healthyWistar rat and the curve 302 is a test of the same rat where the tonguewas treated with a known carcinogen and cancer was allowed to develop.The diagnosis of cancer follows a series of steps;

In a first step, the initial rate of change of voltage "v" (hereinaftercalled the voltage sweep rate) and the distance "d" between theexcitation electrode 100 and the measuring electrode 99 are assumed. Atest run is performed to obtain the current versus voltage graph similarto that of FIG. 7. Numerical data is obtained from the test run andsubstituted into the following equation to obtain a new voltage sweeprate, and a new electrode spacing amongst other parameters. ##EQU1##

where I(B) is the current of the function B and B=E-E_(TPV). E theelectric field strength, E_(TPV) is the electric field at the thresholdpolarisation voltage;

"d" the distance between electrodes:

"υ" the voltage sweep rate in V/S:

"R" the total resistance of the specimen;

τ_(o) is the domain relaxation time constant and τ=τ_(o) exp[U/k T],where U is the activation energy and T the absolute temperature:

"A" is the constant of "softness" which is inversely proportional to thepiezomodulus of the polarising unit (domain, cell, etc.).

The test is then set up to the new voltage sweep, the new electrodespacing and the other parameters, to be run again. This first step isrepeated until all of the parameters in the above equations converge totheir correct values which are determined when the values stop changingsubstantially after each iteration. Finally, a test run with the correctvalues is performed and the threshold polarisation voltage relating toeach domain group, indicated on the curves in FIG. 7 by the localmaxima, is obtained. On these curves a local maxima or humps of a domaingroup having a threshold polarisation voltage of less than 1 volt isindicative of some abnormality.

In a second step the relative permittivity (FIG. 8) and the dissipationfactor (FIG. 9) is obtained as a function of the frequency of theapplied voltage. In FIGS. 8 and 9 the curve marked 201 is the result ofthe measurements of a healthy Wistar rat, the curve marked 202 is theresult of a Wistar rat with an ulcer and curve 203 is a Wistar rat withcancer which is indicated by the local maxima or hump 204 in the curve.

In a third step the specimen is excited or charged to the thresholdpolarisation voltage for each domain independently and allowed todischarge. During this discharge cycle measurements of the dischargecurrent versus time are obtained and analysed to reveal a relaxationtime for each domain. A computer software program designed to analysethe relaxation times for each domain is based on the evaluation of thefollowing equations:

    I(t)=I.sub.o +I.sub.1 exp[-(t/τ.sub.1)]+I.sub.2 exp[-(t/τ.sub.2)]+. . . +I.sub.n exp[-(t/τ.sub.n)],

where I_(n) is the current amplitude and τ_(n) is the relaxation timeconstant for the n^(th) polarised domain group. The computer softwareprogram cross-checks the results of the relaxation time constants by aFourier analysis (as an example of the Fourier analysis the dissipationfactor D for the thigh muscle of a Wistar rat, see FIG. 12) based on theequation: ##EQU2## where C_(n) is the capacitance of the n^(th) domaingroup, which is related to the current amplitude I_(n) and the appliedvoltage "V" by C_(n) =I_(n) τ_(n) /V. C.sub.∝ is the sum of thecapacitance of each domain group and ω is the angular frequency (2πf)for "f" the frequency of the applied voltage "V". D(ω) is the energydissipation as a function of the angular frequency.

The experimental determination of the natural frequency of each domainand hence the relaxation time constants, is obtained by the computersoftware program IEEE 488 from Intel via the measured parameters of thedielectric permittivity and frequency on the basis of the followingequations:

    ε(ω)=ε.sub.r1 (ω)+. . . +ε.sub.rn (ω)+ε.sub.i1 (ω)+. . . +ε.sub.in (ω)

where ##EQU3##

L. C and R are electrical parameters of the equivalent circuit (FIG. 6)correspond to the electromechanical coupling (piezoelectric like) withinthe domains in living cell cytoplasm or between living cells inorganisms. In the equations above: ##EQU4##

The natural frequency is referred to the inductance "L_(in) ",capacitance "C_(in) " and resistance "R_(in) " interrelation of thedomain following the equivalent electrical circuit in FIG. 6. Theresistivity "R" relates to the resistance of each domain group, and δrelates to the piezoelectric constant. The relative permittivity "ε(ω)"is described in the above equation as a function of the angularfrequency, noting that in the equations, the subscript "n" relates tothe n^(th) domain group.

FIG. 10 is an example of a discharge current versus time curve for thedomain structure in a Wistar rat thigh muscle, however, at the top righthand corner of the figure is a table of relaxation time constants, withcorresponding current values and "Q" or charge values for each of fourdomain structures of the cytoplasm. If the "Q"-values of the dischargeprocesses sum up to give the corresponding value calculated from theinput polarisation current, then the test has been successful and therelaxation times of each domain structure correctly reflect thedielectric characteristics of the specimen. These relaxation times arethen compared to average relaxation times for a healthy specimen,similar to the table in FIG. 11. If relaxation time constants of theWistar rat of FIG. 10 are far removed from the values indicated by thetable in FIG. 11 then we can surmise with very good probability thatthere is an abnormality. The abnormality in this case for the Wistar ratof FIG. 10 was cancer.

The third step of the diagnostic procedure is performed within a periodless than or equal to the relaxation time period for the domain andpreferably within the time frame before any substantial change to theintracellular morphology of the cells of the specimen under test. In thepreferred embodiment of the present invention all three steps would beachieved in a few relaxation time cycles.

The DDA as hereinbefore described in the embodiments make possible therecording of the dielectric parameters of tissue samples with minimalinvasion. As the domains in cytoplasm are vulnerable to spontaneousordering, rearrangements or disruption by slight changes, for example bytemperature, the simultaneous measurements of parameters make possiblethe analysis of these changes with reference to the same intracellularmorphology.

FIGS. 13(a)-(f) relate to the changes of polarisation in the tonguetissue with a change in temperature and FIG. 13(f) shows a comparison ofthe minute energies required during heating below 41° C.

FIG. 13(e) illustrates the irreversible process that occurs to thedielectric parameters and hence to living tissue (in this case tonguetissue of a Wistar rat), before and after heating the tissue totemperatures above 42.5 degrees Celsius. The process of heating thetissue above a certain temperature "cooks" the tissue. This "cooking"process changes the state of the dielectric parameters of the tissue,compared to the tissue undergoing chemical "fixation" (chemicals such asKryofix are generally used for optical studies of cellular morphology)which preserves the tissue. These changes in the dielectric parametersare shown in part in FIG. 14.

FIG. 14 is a table showing some dielectric parameters of various tissuesamples of rat organs, averaged over two rats, and a comparison of theseparameters for fresh or "fixated" tissue.

Industrial Applicability

The dielectric diagnostic analyser (DDA) as described in the embodimentsof the present invention also provides a non-invasive, or at leastminimally invasive, technique to diagnose changes in the fine structurein cell cytoplasm with respect to the complexity of chemical context.cellular packing, disease transformation and reveal the action ofpreservation (e.g., Kryofix) and staining (e.g., Haematoxylin) ontissue.

It will be appreciated by a person skilled in the relevant art that thismethod of diagnostic testing can be applied to any specimen or substancewhere a domain type structure within cells can be defined including anyMaxwell-Wagner system. To study ultrafine structure and intracellularkinetic parameters of cells, including the cell cytoplasm, tissue,organs, the body's metabolic processes, the detection of disease anddisease transformation at the onset of said disease including thedifferentiation of diseases having or not having a viral origin.

The method herein described provides a diagnostic tool which can beadapted to imaging techniques, similar to medical imaging. Thisdiagnostic method and apparatus can be adapted to animals in animalhusbandry, plants in agriculture, environmental diagnostics of bacteriaand algae in waterways and to chemical analysis of effluent amongstother fields of use. Typically the diagnostic method hereinbeforedescribed is well suited to the analysis of the presence or absence oftoxins and other chemicals in specimens such as dairy products,vegetables, meat, fruit, fish, grains, oils, seeds and stock feedproducts, soil, water as well as viral diseases in plants, animals orhuman bodies.

It will be appreciated by persons skilled in the art that numerousvariations and/or modifications may be made to the invention as shown inthe specific embodiments without departing from the spirit or scope ofthe invention as broadly described. The present embodiments are,therefore, to be considered in all respects as illustrative and notrestrictive.

We claim:
 1. An apparatus adapted to perform diagnostic analysis of aspecimen having at least one domain group as hereinbefore defined, theapparatus comprising:excitation generating means to generate apredetermined excitation signal; measuring means to measure a responsesignal of the specimen to the predetermined excitation signal; electrodemeans for transmitting and receiving the predetermined excitation signaland response signal of the specimen, respectively; analysing meansarranged to analyse said response signal; and switching means adapted toswitch the electrode means between the measuring means, and excitationgenerating means, in a time period less than a polarization relaxationtime period of the at least one domain group in the specimen, theswitching means being controlled by the analysing means to switch theelectrode means between the excitation means and the measuring means attimes less than the smallest relaxation time to be measured.
 2. Anapparatus as claimed in claim 1 wherein the excitation generating meansis the source of the predetermined excitation signal.
 3. An apparatus asclaimed in claim 2 wherein the excitation generating means is anelectrometer.
 4. An apparatus as claimed in claim 2 wherein theexcitation generating means is a frequency bridge.
 5. An apparatus asclaimed in claim 1 wherein the analysing means comprises an electroniccomputer.
 6. An apparatus as claimed in claim 1, wherein the electrodemeans is a suction cup electrode, a pinch electrode or athermocontrolled electrode.
 7. A method of diagnostic analysiscomprising:applying a predetermined first excitation signal to aspecimen having at least one domain group, as hereinbefore defined, soas to elicit a response from the domain group; analysing the responsefrom the domain group to determine the maximum response of each domaingroup; comparing said maximum response to a maximum response of acontrol specimen; applying a second excitation signal corresponding to asignal value at, or near, the point of maximum response of each domaingroup to elecit a further response in each domain group; and detectingthe variation and length of said further response upon removal of thesecond excitation signal.
 8. A method as claimed in claim 7 wherein thefirst excitation signal is a ramp function voltage sweep or a time rateof change of voltage, and the response from the domain, in the domaingroup of the system, is measured as a change in a current flow throughthe system over time.
 9. A method as claimed in claim 7 wherein thefirst excitation signal is a frequency dependent applied voltage and theresponse from the domains is measured so as to allow the determinationof dielectric permittivity, and dissipation energy, of each domaingroup.
 10. A method as claimed in claim 7 wherein the second excitationsignal is applied in the absences of the first excitation signal.
 11. Amethod as claimed in claim 7 or 10 wherein the further response ismeasured upon removal of the second excitation signal while each domainis relaxing to its natural state.
 12. A method as claimed in any one ofthe claims 7, 10 and 11 wherein the detecting of the variation andlength the further response occurs within the time in which the domainsin each group relax to the state they were in before the secondexcitation signal was applied.
 13. A method as claimed in claim 7,wherein the specimen is of human cells or tissue.
 14. A method asclaimed in claim 7, wherein the specimen is other than of human cells ortissue.